High chimera rate: +80%

[Mx-nene]

Hi.

I am doing microbiome Research on chicken stool.

I have ran to a problem with my analysis. I have ran DADA2 on Illumina MiSeq 2x150bp 16S rRNA sequence data, and obtained a chimera percentage of 83% which is problematic. I did use the primers 341f (CCTACGGGNGGCWGCAG) and 785r (GACTACHVGGGTATCTAATCC) for trimming but I have the 83% chimeric sequences.

Could you please provide you valuable insight on such a a problem, because, I should be submitting my dissertation and article publications soon and this error is hindering me.

I hope my request receives favourable consideration.

[benjjneb]

Is this the percent of the ASVs in the data? Or the percent of the reads in the data? Quoting the dada2 tutorial section on chimera removal:

Considerations for your own data: Most of your reads should remain after chimera removal (it is not uncommon for a majority of sequence variants to be removed though).

[Mx-nene]

When looking for the abundance of merged reads, I'm assuming it's the ASVs in the data, 83% are considered as chimeras.

The R code was:

sum(seqtab.nochim) / sum(seqtab)

[benjjneb]

If the result of sum(seqtab.nochim)/sum(seqtab) is ~0.83, it means that 83% of your reads are non-chimeras (so 17% are being removed as chimeras).

[Mx-nene]

That's perfect.

Thank you very much, Dr Callahan.